P11.29 Development of ex vivo models for deeper insights into the biology and therapeutic targeting of tumor microtube networks in gliomas

Abstract BACKGROUND The formation of multicellular networks via thin cellular protrusions named tumor microtubes (TMs) emerged as a novel mechanism of therapy resistance in malignant glioma. TMs are also involved in tumor cell invasion and growth. Within these tumor cell networks, connected tumor cells communicate via intercellular calcium waves (ICWs). Only few molecular drivers of TMs (Gap43, Ttyh1, Connexin 43) have been identified until now. Furthermore, the molecular mechanisms underlying ICWs as well as their specific biological role in glioma remains to be elucidated. A better understanding of the biology and the identification of molecular key drivers is essential for the development of drugs targeting TM formation and function. MATERIAL AND METHODS For this purpose, we have developed novel ex vivo models that not only provide insights into TM biology but further allow medium throughput drug screening. As classical response parameters such as the inhibition of cell growth or cytotoxicity do not necessarily correlate with effects on TM formation or function, a morphometrical approach employing laser scanning microscopy and machine-learning based image analysis tools is used. The application of fluorescent probes and genetic fluorescent reporter systems provides novel longitudinal insights into cytoskeletal dynamics, the role and exchange of organelles such as mitochondria, mechanisms of homeostasis within tumor cell networks (e.g. redox homeostasis) and ICWs in live cells. In addition to 2D glioma cell and co-culture models we have developed a fully human and mature brain organoid model. Here, complex 3D tumor cell networks corresponding to the morphology and exhibiting calcium communication patterns observed in our mouse model can be established and studied ex vivo. Furthermore, with these models not only the role of the brain microenvironment on TM formation but also direct interactions of glioma cells with neurons and glial cells as well as drug effects such as cytotoxicity on these brain cells can be investigated ex vivo. CONCLUSION In summary, novel tumor models enable further insights into TM biology and hence provide the basis for development of TM- and network disrupting drugs. First results of this screening opportunity will be presented.